Cholera is a severe diarrheal disease that affects approximately 3 million individuals per year worldwide (mainly in less developed countries). It is caused by consuming food or drinking water contaminated with the microorganism Vibrio cholerae. When the organism is ingested, it has the ability to colonize the intestinal tract.
In the small intestine, V. cholerae attaches to the intestinal mucosa and releases exotoxins, the most important being cholera toxin (CT), which act on mucosal cells 1-4!. The action of CT on intestinal cells induces fluid secretion and increased permeability of electrolytes into the small intestine resulting in severe diarrhea and electrolyte imbalance.
Two other toxins are also produced by V. cholerae. They include zona occludens toxin (Zot) which disrupts tight junctions between cells, and accessory cholera enterotoxin (Ace), which causes diarrhea in animals. The role of these two toxins in the overall pathogenesis of the disease remains unclear.
An additional cytolytic toxin is produced by O1 El Tor and O139 serotypes of V. cholerae. This toxin has a hemolytic and cytotoxic activity which appears to play a role in the pathogenesis of cholera.
Mortality rates are high for infants and children that are inflicted with cholera. The current method of treatment for cholera is to replace fluids and restore electrolyte balance.
Not all strains of V. cholerae are responsible for causing disease. The disease causing strains belong to the O1 serotype which includes the classical and the El Tor biotypes. All other serotypes except for one are thought to be nonvirulent or capable of causing only minor diarrhea. The only non O1 strain of V. cholerae that has been shown to cause full-blown cholera was identified two years ago. It belongs to the O139 serotype. This serotype has been identified as the causal agent for recent outbreaks of cholera in Asia. It produces all the virulence factors (including CT) associated with the O1 serotypes of V. cholerae.
The virulence factors most important for causing disease are the toxin coregulated pili (Tcp) which allow V. cholerae to colonize the small intestine. Although the host cell receptor has yet to be identified for pili, there is some indirect evidence which suggests that a carbohydrate may be involved. This evidence is based on the finding that individuals who have the O blood group are more susceptible to severe cases of cholera while people who are AB blood group positive tend to be somewhat resistant toward the disease. One possible explanation for this finding is that the pili found on V. cholerae may use the O blood group oligosaccharide structure for colonization of the small intestine, thus rendering individuals with the O blood group more susceptible to disease.
CT is the virulence factor most responsible for the symptoms of the disease. CT possesses an enzymatic activity which elevates the levels of cyclic AMP (cAMP) in host cells. The increase in cAMP levels alters the ion transport systems within cells thus affecting the osmotic balance within the intestine that leads to diarrhea. CT utilizes the ganglioside GM1 (.beta.Gal(1-3).beta.GalNAc(1-4).alpha.NeuAc(2-3)!.beta.Gal(1-4).beta.Glc -ceramide) to bind to host cell receptors.
Cholera toxin (CT) has been shown to bind to several derivatives of the ganglioside GM1 where the carboxyl group of sialic acid had been modified to form a number of C(1) amides 5!. The structure of these compounds is: .beta.Gal(1-3) .beta.GalNAc(1-4).alpha.NeuAcR(2-3)! .beta.Gal(1-4).beta.Glc-ceramide, where R is selected from the group consisting of amide, methylamide, ethylamide, propylamide, and benzylamide of sialic acid.
Other derivatives of GM1 that were shown to bind CT include 6!: .beta.Gal(1-3) .beta.GalNH2(1-4).alpha.Neu-NH2(2-3)! .beta.Gal(1-4).beta.Glc-ceramide; .beta.Gal(1-3) .beta.GalNAc(1-4).alpha.NeuAcR(2-3)! .beta.Gal(1-4).beta.Glc-ceramide, where R is the methyl ester of sialic acid; .beta.Gal(1-3).beta.GalNAc(1-4).alpha.(C7)NeuAc(2-3)! .beta.Gal(1-4).beta.Glc-ceramide; and .beta.Gal(1-3) .beta.GalNAc(1-4).alpha.NeuAcR(2-3)! .beta.Gal(1-4).beta.Glc-ceramide, where R is ethanolamineamide.
Other gangliosides which have been shown to bind CT include6,7!: GM2 (.beta.GalNAc(1-4).alpha.NeuAc(2-3)! .beta.Gal(1-4).beta.Glc-ceramide) and GD1b (.beta.Gal(1-3) .beta.GalNAc(1-4).alpha.NeuAc(2-3).alpha.NeuAc(2-3)! .beta.Gal(1-4).beta.Glc-ceramide.
In addition, highly purified CT preparations have been obtained using lyso GM1 ganglioside or galactose affinity columns 8-10!.
With respect to methods of diagnosis of the presence of CT in a sample, one method for detecting Vibrio cholerae in a sample is to culture the sample. The disadvantages of this method include the length of time required and interference by non-pathogenic, i.e., non-toxin producing, V. cholerae strains. Other methods involve the use of specific antisera or monoclonal antibodies.
In view of the above, there is a need for a compound which would treat cholera. A preferred compound would be administered noninvasively, such as orally, and would specifically remove toxin and/or organisms from the intestinal tract.